In the past decade about 15 TSG(s) were identified and cloned. With the gene in hand the predictions of Knudson's foundational theory of human carcinogenesis were verified and mutated cancer causing TSGs became the paradigm of genetic etiology and pathogenesis of inherited and common cancers. The ultimate goals of our research were (1) to complete the isolation and characterization of the VHL TSG and to begin to understand its biochemical function; (2) and to identify one or more TSGs from the 3p21-p22 region, involved in the causation of lung cancer(s). Our major accomplishments this year are: (1) identification and analysis of the promoter of the VHL gene; identification of the long 3' UTR and construction of a VHL intronless gene; construction of a set of VHL sense and antisense minigenes in diverse expression vectors (including episomal and retroviral) for ex vivo/in vivo delivery; establishment of renal carcinoma (RCA) cell lines overexpressing VHL transgenes and pVHL/vhilin. (2) The positioning of the SCLC TSG within the telomeric part of a 700 kb cosmid - P1 clone contig spanning the candidate region as defined by overlapping deletions and allelotyping. A number of potential TSG mapped by us and others to 3p21-p22 namely, GNAI-2, GNAT-1, Wint 5, beta catenin, TGFbetaRII, and MLH1, were excluded from this candidate region. To date 12 new genes were isolated, partial or complete cDNAs sequenced, and their candidacy is being evaluated by mutational analysis (SSCP of exons or RT PCR followed by sequencing) and expression in SCLC cell lines.